In Message-ID: <Jan.Svensson.23.2FF800C4 at mikrob.slu.se>
Jan.Svensson at mikrob.slu.se (Jan Svensson) wrote:
>I have a question about the Ni-NTA resin (qiagen/diagen).
>In The QIAexpressionist (booklet fronm qiagen 2 ed 1992) they state that about
>the Ni-NTA resin "re-use of the Ni-NTA resin..should only be performed with
>identical recombinant proteins..maximum of 3-5 runs per column"
>In the booklet there is a regenaration procedure described containing 17 steps.
>Isn't it possible to use the column for more than one protein as long the
>column is intact?
>Can't the regeneration be done in an easier way (less than 17 steps), just
>strip it with EDTA and recharge the column with NiSo4 ?
Hi Jan,
Ni-NTA resin can probably be reused after stripping the column with EDTA
and recharging it with Ni ions. However, this will give you no guarantee
that you have removed all the residual proteins and you risk contamination
of your next prep. Also, the resin properties will progressively
deteriorate as the level of residual contamination increases.
The long washing procedure we describe is tedious, we know, but it is the
only way to be absolutely sure you have removed all contaminants (both
protein and non-protein) that may be interacting with the resin. Most users
find that it is safer, more convenient, and more cost-effective (unless you
are using extremely high resin volumes) to use fresh Ni-NTA resin where
possible.
Also, you should note that Ni-NTA resin will bind 5Ð10 mg of protein per ml
of resin, and that the purification procedure is more efficient if you use
less resin than you actually need, and overload it somewhat. The QIAGEN
Technical Service Department has extensive references for this system, and
can help you optimize your purification procedure by using less Ni-NTA
resin.
QIAGEN Technical Service Department
800-426-8157
--
QIAGEN 800-362-7737
Temporary Email Address qiagen at kaiwan.com